dc.contributor.author | Papple, Alex G. | |
dc.date.accessioned | 2016-06-23T11:41:06Z | |
dc.date.available | 2016-06-23T11:41:06Z | |
dc.date.issued | 2015-10-30 | |
dc.date.submitted | 2016-06-15T14:52:51.748Z | |
dc.identifier.citation | Papple, A.G., 2015: INVESTIGATING DUAL-TARGETED DOMAIN ANTIBODY FUSION PROTEINS IN A CANCER MODEL, Queen Mary University of London. | en_US |
dc.identifier.uri | http://qmro.qmul.ac.uk/xmlui/handle/123456789/13033 | |
dc.description | PhD Thesis | en_US |
dc.description.abstract | Developing efficacious, highly specific therapeutics is an ongoing challenge.
The domain antibody (dAb) fragment platform has shown promise targeting a dAbfused
therapeutic moiety to a specific tissue. This investigation sought to conclude on
the optimum binding affinity combination to maximise tissue-targeting specificity and
localised therapeutic potency by using a panel of dAb fusion proteins with varying
affinity combinations of asialoglycoprotein receptor (ASGPR)-targeting dAb mutants
and human interferon (hIFN) mutants – hIFN-ASGPRdAbs.
The hIFN-ASGPRdAbs were engineered, expressed, 1,4,7-
triazacyclononanetriacetic acid (NOTA)-conjugated and purified with binding affinities
and potencies characterised. Gallium-68 radiolabelling of the NOTA-hIFNASGPRdAbs
was optimised using a high activity fraction in a 10 minute reaction at
ambient temperature in pH4.4 sodium acetate. HepG2 xenograft mouse models
were injected with 68Ga-NOTA-hIFN-dAbs for biodistribution analysis and
PET/CT imaging, followed by mRNA expression analysis of the xenograft tissue.
ASGPR binding affinities of the NOTA-hIFN-ASGPRdAbs ranged from
0.73pM to ~528nM, and their hIFN potency ranged from 6.79pM to ~3.87nM.
Targeting of HepG2 cells was driven by the dAb-mediated ASGPR targeting. Ga-68
radiolabelling efficiencies up to 98.4% and specific activities up to 2.47MBq/μg were
achieved. Xenograft uptake was significantly increased through dAb-mediated
ASGPR targeting compared to the non-ASGPR targeted control 68Ga-NOTA-hIFNCTRLdAb,
achieving 2.42%ID/g vs. 0.68%ID/g, respectively. A trend of increasing
xenograft uptake correlated with increasing ASGPR binding affinity for the panel of
mutants, in spite of very high murine liver uptake. Increased xenograft uptake also
correlated with a more potent hIFN-mediated anti-proliferative mRNA response in
the xenograft tissue. A dAb with a mid-range affinity for ASGPR and maximum
affinity hIFN was concluded to be the optimum combination.
These results showed that the efficacy of a dAb fusion protein can be
influenced by the two intrinsic binding affinities. Hence, dAb-mediated tissue
targeting of a fused therapeutic and engineered affinity synergism strategies may hold
the key to novel, selectively cytotoxic biopharmaceutical drugs. | |
dc.description.sponsorship | Queen Mary University of London,
GlaxoSmithKline (GSK) and the EPRSC. | en_US |
dc.language.iso | en | en_US |
dc.publisher | Queen Mary University of London | en_US |
dc.subject | Medicine | en_US |
dc.subject | therapeutics | en_US |
dc.subject | cytotoxic biopharmaceutical drugs. | en_US |
dc.title | INVESTIGATING DUAL-TARGETED DOMAIN ANTIBODY FUSION PROTEINS IN A CANCER MODEL | en_US |
dc.type | Thesis | en_US |
dc.rights.holder | The copyright of this thesis rests with the author and no quotation from it or information derived from it may be published without the prior written consent of the author | |